Research Summary: The KRAS proto-oncogene is one of the most critical genes in cancer, yet it has also proven to be among the most elusive. Nearly all (98%) of KRAS missense mutations occur in codons 12 or 13, which leads to constitutive KRAS activation and promotion of numerous cancer hallmarks. Although kinase inhibitors have revolutionized treatment of some subsets of cancers driven by other molecular aberrations, the lack of such success in mutant KRAS-driven cancers has led KRAS itself being widely regarded as ?undruggable?. Although there are G12C inhibitors currently being developed, these therapies are limited to only one KRAS mutation, accounting for only ~12% of KRAS mutations, and therefore limited in therapeutic scope. We have established a C Corporation, EnFuego Therapeutics, Inc. to address the growing number of ?undruggable? targets in cancer by using RNA interference (RNAi)-based therapeutics. RNAi is attractive because it enables silencing of cancer targets that cannot be inhibited using conventional approaches. We have several lines of evidence that our lead drug, EFTX- 001, potently silences the majority of missense mutations and spares the WT sequence. Using an FDA-approved lipid nanoparticle (LNP) delivery system, we have already found impressive therapeutic potential for delivering EFTX-001 in lung cancer models with no detectable toxicity. Taken together, key questions arise, such as: 1) Which KRAS missense mutations are most potently targeted by EFTX-001? 2) What toxic effects does LNP- EFTX-001 have on healthy adult tissue (e.g. liver, kidney and bone marrow)? 3) What is the therapeutic index of LNP-EFTX-001 in KRAS-dependent cancer models? We hypothesize that LNP-EFTX-001 will potently silence multiple clinically-relevant KRAS mutations in lung and colon cancers while sparing the wild-type sequence; consequently, inhibiting tumor progression with a highly satisfactory toxicity profile. The objectives of this proposal are: 1) to define how potently EFTX-001 targets the most common KRAS mutations while sparing the WT sequence, 2) to evaluate the effects of EFTX-001 on MAP kinase signaling and cell viability, 3) use in vitro and in vivo models to determine the therapeutic index of silencing mutant versus WT KRAS in cancer and adult tissues, respectively.